Biomedical subjects

Michael Frotscher

Publications and source records attributed to Michael Frotscher.

49 records · Page 3Linked to original sources

PubMed2003

Comparison of commissural sprouting in the mouse and rat fascia dentata after entorhinal cortex lesion.

Reactive axonal sprouting occurs in the fascia dentata after entorhinal cortex lesion. This sprouting process has been described extensively in the rat, and plasticity-associated molecules have been identified that might be involved in its regulation. To demonstrate causal relationships between these candidate molecules and the axonal reorganization process, it is reasonable to analyze knockout and transgenic animals after entorhinal cortex lesion, and because gene knockouts are primarily generated in mice, it is necessary to characterize the sprouting response after entorhinal cortex lesion in this species. In the present study, Phaseolus vulgaris-leucoagglutinin (PHAL) tracing was used to analyze the commissural projection to the inner molecular layer in mice with longstanding entorhinal lesions. Because the commissural projection to the fascia dentata is neurochemically heterogeneous, PHAL tracing was combined with immunocytochemistry for calretinin, a marker for commissural/associational mossy cell axons. Using both techniques singly as well as in combination (double-immunofluorescence) at the light or electron microscopic level, it could be shown that in response to entorhinal lesion mossy cell axons leave the main commissural fiber plexus, invade the denervated middle molecular layer, and form asymmetric synapses within the denervated zone. Thus, the commissural sprouting response in mice has a considerable translaminar component. This is in contrast to the layer-specific commissural sprouting observed in rats, in which the overwhelming majority of mossy cell axons remain within their home territory. These data demonstrate an important species difference in the commissural/associational sprouting response between rats and mice that needs to be taken into account in future studies.

Animals↗

PubMed2003

Reelin controls granule cell migration in the dentate gyrus by acting on the radial glial scaffold.

Reelin, synthesized and secreted by Cajal-Retzius (CR) cells in the marginal zone of the cortex, is an extracellular matrix protein important for the development of cortical layers. In reeler mutant mice lacking Reelin, there are severe malformations of neocortical and hippocampal lamination. It has been assumed that Reelin acts as a stop signal for migrating neurons. Here we show, by using the dentate gyrus as a model in in vivo studies and in vitro assays, that Reelin exerts its effects, at least in part, by acting on the radial glial scaffold required for neuronal migration. Migration defects of dentate granule cells, reminiscent of those seen in reeler mutants, are observed in tissue from patients with temporal lobe epilepsy (TLE). The extent of granule cell dispersion in TLE was found to be inversely correlated with the number of reelin mRNA synthesizing CR cells and reelin mRNA expression as revealed in quantitative RT-PCR studies. These findings show that the Reelin signaling pathway is essential for the correct positioning of human hippocampal neurons and that a Reelin deficiency is involved in the pathological changes associated with epilepsy.

Adult↗

PubMed2003

Reelin signaling directly affects radial glia morphology and biochemical maturation.

Radial glial cells are characterized, besides their astroglial properties, by long radial processes extending from the ventricular zone to the pial surface, a crucial feature for the radial migration of neurons. The molecular signals that regulate this characteristic morphology, however, are largely unknown. We show an important role of the secreted molecule reelin for the establishment of radial glia processes. We describe a significant reduction in ventricular zone cells with long radial processes in the absence of reelin in the cortex of reeler mutant mice. These defects were correlated to a decrease in the content of brain lipid-binding protein (Blbp) and were detected exclusively in the cerebral cortex, but not in the basal ganglia of reeler mice. Conversely, reelin addition in vitro increased the Blbp content and process extension of radial glia from the cortex, but not the basal ganglia. Isolation of radial glia by fluorescent-activated cell sorting showed that these effects are due to direct signaling of reelin to radial glial cells. We could further demonstrate that this signaling requires Dab1, as the increase in Blbp upon reelin addition failed to occur in Dab1-/- mice. Taken together, these results unravel a novel role of reelin signaling to radial glial cells that is crucial for the regulation of their Blbp content and characteristic morphology in a region-specific manner.

Animals↗

PubMed2002

Three-dimensional reconstruction of a calyx of Held and its postsynaptic principal neuron in the medial nucleus of the trapezoid body.

The three-dimensional morphology of the axosomatic synaptic structures between a calyx of Held and a principal neuron in the medial nucleus of the trapezoid body (MNTB) in the brainstem of young postnatal day 9 rats was reconstructed from serial ultrathin sections. In the apposition zone between the calyx and the principal neuron two types of membrane specializations were identified: synaptic contacts (SCs) with active zones (AZs) and their associated postsynaptic densities (PSDs) constituted approximately 35% (n = 554) of the specializations; the remaining 65% (n = 1010) were puncta adherentia (PA). Synaptic contacts comprised approximately 5% of the apposition area of presynaptic and postsynaptic membranes. A SC had an average area of 0.100 microm(2), and the nearest neighbors were separated, on average, by 0.59 microm. Approximately one-half of the synaptic vesicles in the calyx were clustered within a distance of 200 nm of the AZ membrane area, a cluster consisting of approximately 60 synaptic vesicles (n = 52 SCs). Approximately two synaptic vesicles per SC were "anatomically docked." Comparing the geometry of the synaptic structure with its previously studied functional properties, we find that during a single presynaptic action potential (AP) (1) approximately 35% of the AZs release a transmitter quantum, (2) the number of SCs and anatomically docked vesicles is comparable with the low estimates of the readily releasable pool (RRP) of quanta, and (3) the broad distribution of PSD areas [coefficient of variation (CV) = 0.9] is likely to contribute to the large variability of miniature EPSC peaks. The geometry of the reconstructed synapse suggests that each of the hundreds of SCs is likely to contribute independently to the size and rising phase of the EPSC during a single AP.

Animals↗

PubMed2002

Laminar distribution of synaptopodin in normal and reeler mouse brain depends on the position of spine-bearing neurons.

Synaptopodin is the first member of a novel class of proline-rich actin-associated proteins. In brain, it is present in the neck of a subset of mature telencephalic spines and is associated closely with the spine apparatus, a Ca(2+) storing organelle within the spine compartment. The characteristic region- and lamina-specific distribution of synaptopodin in rat brain suggested that the distribution pattern of synaptopodin depends on the cytoarchitectonic arrangement of spine-bearing neurons. To test this hypothesis, synaptopodin was studied in the cortex, striatum, and hippocampus of normal and reeler mice, in which developmental cell migration defects have disrupted the normal array of cells. In situ hybridization histochemistry as well as light- and electron microscopic immunocytochemistry were used. In brain of normal mice, the pattern of synaptopodin mRNA-expressing cells corresponds to that of spine-bearing neurons and synaptopodin protein is found in a region- and lamina-specific distribution pattern. It is specifically sorted to the spine neck where it is associated closely with the spine apparatus. In brain of reeler mice, the pattern of synaptopodin mRNA-expressing cells corresponds to that of the abnormally positioned spine-bearing neurons and the region- and lamina-specific distribution pattern is absent or altered. Nevertheless, synaptopodin was specifically sorted to the spine neck, as in controls. These data demonstrate that the light microscopic distribution pattern of synaptopodin protein depends on the position and orientation of the spine-bearing neurons. The intracellular sorting process, however, is independent of positional cues.

Animals↗

PubMed2002

Anterograde tracing of human hippocampus in vitro-a neuroanatomical tract tracing technique for the analysis of local fiber tracts in human brain.

Hippocampal slices were obtained from hippocampi of patients with temporal lobe epilepsy or from patients with mass lesions located in the temporal lobe. Hippocampal slices were kept alive in a slice chamber and the anterograde tracer neurobiotin was iontophoretically injected into the granule cell layer of the dentate gyrus. Single injections resulted in the labeling of small groups of granule cells. The axonal arbor of these cells could be partially reconstructed and single mossy fibers could be followed from the soma to the inner molecular layer of the sclerotic dentate gyrus. Electron microscopy revealed asymmetric mossy fiber synapses on spiny neurons in the inner molecular layer, presumably granule cells. These data demonstrate that in vitro anterograde tracing can be employed to study the local connectivity of the human brain at the light and electron microscopic level.

Biotin↗

PubMed2002

Synaptic vesicle protein synaptoporin is differently expressed by subpopulations of mouse hippocampal neurons.

In the hippocampus, the synaptic vesicle protein synaptoporin (SPO) has been reported to be exclusively enriched in the granule cell axons, the mossy fibers. In this study, we show that in adult rats and mice SPO immunoreactivity (IR) is also detectable in strata oriens, radiatum, and lacunosum-moleculare of CA1-CA3, as well as perisomatically in the hippocampus proper and fascia dentata. In situ hybridization confirmed that SPO mRNA was present in granule cells and CA3 pyramidal cells but not in CA1 pyramidal cells. Importantly, cells scattered throughout the hippocampal layers resembling the distribution of interneurons were found to synthesize high amounts of SPO mRNA, too. Thus, these findings indicate that SPO expression in the hippocampus was underestimated until now. Moreover, double-labeling immunohistochemistry and confocal microscopy revealed selective colocalization of SPO and glutamate decarboxylase (GAD 65), a marker for gamma-aminobutyric acid (GABA)ergic terminals. To identify SPO expressing interneurons, in situ hybridization was combined with immunocytochemistry against parvalbumin (PV), calbindin (CB), calretinin (CR), cholecystokinin (CCK), and vasoactive intestinal polypeptide (VIP). We found that SPO transcripts were differentially expressed by various interneuron subpopulations in the hippocampus of C57Bl/6 mice (PV 44.2%, CB 46.3%, CR 19.3%, CCK 38.6%, VIP 59.9%). Immunoelectron microscopy for SPO labeled synaptic vesicle profiles in distinct symmetric and asymmetric synapses. In conclusion, our data demonstrate that hippocampal principal cells and interneurons display a variety of synaptic vesicles that are likely to contribute to the functional characteristics of their output synapses.

Animals↗

PubMed2002

Reelin, Disabled 1, and beta 1 integrins are required for the formation of the radial glial scaffold in the hippocampus.

The extracellular matrix molecule Reelin is required for the correct positioning of neurons during the development of the forebrain. However, the mechanism of Reelin action on neuronal migration is poorly understood. Reelin is assumed to act on neurons directly, but it may also affect the differentiation of glial cells necessary for neuronal migration. Here we show that a regular glial scaffold fails to form in vivo in the dentate gyrus of mice deficient of Reelin or Disabled 1, a neuronal adaptor protein in the Reelin signaling pathway. A subset of these defects is observed in mice that lack beta(1)-class integrins, known to bind Reelin. Moreover, recombinant Reelin induced branching of glial processes in vitro. Our data suggest that Reelin affects glial differentiation via Disabled 1 and beta(1)-class integrin-dependent signaling pathways.

Animals↗

PubMed2002

Fast synaptic inhibition promotes synchronized gamma oscillations in hippocampal interneuron networks.

Networks of GABAergic interneurons are of critical importance for the generation of gamma frequency oscillations in the brain. To examine the underlying synaptic mechanisms, we made paired recordings from "basket cells" (BCs) in different subfields of hippocampal slices, using transgenic mice that express enhanced green fluorescent protein (EGFP) under the control of the parvalbumin promoter. Unitary inhibitory postsynaptic currents (IPSCs) showed large amplitude and fast time course with mean amplitude-weighted decay time constants of 2.5, 1.2, and 1.8 ms in the dentate gyrus, and the cornu ammonis area 3 (CA3) and 1 (CA1), respectively (33-34 degrees C). The decay of unitary IPSCs at BC-BC synapses was significantly faster than that at BC-principal cell synapses, indicating target cell-specific differences in IPSC kinetics. In addition, electrical coupling was found in a subset of BC-BC pairs. To examine whether an interneuron network with fast inhibitory synapses can act as a gamma frequency oscillator, we developed an interneuron network model based on experimentally determined properties. In comparison to previous interneuron network models, our model was able to generate oscillatory activity with higher coherence over a broad range of frequencies (20-110 Hz). In this model, high coherence and flexibility in frequency control emerge from the combination of synaptic properties, network structure, and electrical coupling.

Animals↗

PubMed2002

Role for reelin in the development of granule cell dispersion in temporal lobe epilepsy.

The reelin signaling pathway plays a crucial role during the development of laminated structures in the mammalian brain. Reelin, which is synthesized and secreted by Cajal-Retzius cells in the marginal zone of the neocortex and hippocampus, is proposed to act as a stop signal for migrating neurons. Here we show that a decreased expression of reelin mRNA by hippocampal Cajal-Retzius cells correlates with the extent of migration defects in the dentate gyrus of patients with temporal lobe epilepsy. These results suggest that reelin is required for normal neuronal lamination in humans, and that deficient reelin expression may be involved in migration defects associated with temporal lobe epilepsy.

Adaptor Proteins, Signal Transducing↗

PubMed2002

Abnormal positioning of granule cells alters afferent fiber distribution in the mouse fascia dentata: morphologic evidence from reeler, apolipoprotein E receptor 2-, and very low density lipoprotein receptor knockout mice.

The fascia dentata of the hippocampal formation is characterized by the nonoverlapping and lamina-specific termination of afferent fibers: entorhinal fibers terminate in the outer molecular layer and commissural/associational fibers terminate in the inner molecular layer. It has been proposed that this fiber lamination depends on the presence of the correct postsynaptic partner at the time of fiber ingrowth during development. Pioneer neurons that guide afferent fibers to their correct layers as well as signals located on granule cells have both been implicated. To study the role of granule cells for the lamina-specific ingrowth of afferents, the cyto- and fiberarchitecture of three mouse mutants (very low density lipoprotein receptor knockout mouse, apolipoprotein E receptor 2 knockout mouse, and reeler mouse) that show different degrees of granule cell migration defects were analyzed. Anterograde tracing with Phaseolus vulgaris-leucoagglutinin was used to visualize the afferent fiber systems, and immunohistochemistry was used to determine the position of their putative target cells. In controls, granule cells are packed in a single layer. This laminar organization is mildly altered in very low density lipoprotein receptor knockout mice, moderately disturbed in apolipoprotein E receptor 2 knockout mice, and severely disrupted in reeler mice. These changes in granule cell distribution are mirrored by the distribution of commissural fibers. In contrast, changes in granule cell distribution do not severely affect the laminar termination of entorhinal fibers. These data provide further evidence for a role of granule cells in the laminar termination of commissural/associational afferents to the fascia dentata.

Afferent Pathways↗

PubMed2002

Complete deletion of the neurotrophin receptor p75NTR leads to long-lasting increases in the number of basal forebrain cholinergic neurons.

Cholinergic neurons innervating cortical structures are among the most affected neuronal populations in Alzheimer's disease. In rodents, they express high levels of the neurotrophin receptor p75NTR. We have analyzed cholinergic septohippocampal neurons of the medial septal nucleus in p75exonIII (partial p75NTR knock-out) and p75exonIV (complete p75NTR knock-out) mice, in their original genetic background and in congenic strains. At postnatal day 15, the p75exonIII mutation leads to a moderate increase (+13%) in these neurons among littermates only after back-crossing in a C57BL/6 background. In contrast, the null p75exonIV mutation, which prevents expression of both the full-length and the shorter p75NTR isoforms, results in a 28% neuronal increase, independent of genetic background. The incomplete nature of the p75NTR mutation used previously, coupled with difficulties in delineating the mouse medial septum and the impact of the genetic background on cell numbers, all contribute to explain previous difficulties in establishing the role of p75NTR in regulating cholinergic neuron numbers in the mouse forebrain.

Acetylcholine↗

PubMed2002

Dentate granule cells in reeler mutants and VLDLR and ApoER2 knockout mice.

We have studied the organization and cellular differentiation of dentate granule cells and their axons, the mossy fibers, in reeler mutant mice lacking reelin and in mutants lacking the reelin receptors very low density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2). We show that granule cells in reeler mice do not form a densely packed granular layer, but are loosely distributed throughout the hilar region. Immunolabeling for calbindin and calretinin revealed that the sharp border between dentate granule cells and hilar mossy cells is completely lost in reeler mice. ApoER2/VLDLR double-knockout mice copy the reeler phenotype. Mice deficient only in VLDLR showed minor alterations of dentate organization; migration defects were more prominent in ApoER2 knockout mice. Tracing of the mossy fibers with Phaseolus vulgaris leukoagglutinin and calbindin immunolabeling revealed an irregular broad projection in reeler mice and ApoER2/VLDLR double knockouts, likely caused by the irregular wide distribution of granule cell somata. Mutants lacking only one of the lipoprotein receptors showed only minor changes in the mossy fiber projection. In all mutants, mossy fibers respected the CA3-CA1 border. Retrograde labeling with DiI showed that malpositioned granule cells also projected as normal to the CA3 region. These results indicate that ( 1 ) reelin signaling via ApoER2 and VLDLR is required for the normal positioning of dentate granule cells and (2) the reelin signaling pathway is not involved in pathfinding and target recognition of granule cell axons.

Animals↗